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. 2000 Aug;38(8):2940-2.
doi: 10.1128/JCM.38.8.2940-2942.2000.

Detection of Paracoccidioides brasiliensis in tissue samples by a nested PCR assay

R Bialek  1 , A Ibricevic, C Aepinus, L K Najvar, A W Fothergill, J Knobloch, J R Graybill
Affiliations

Detection of Paracoccidioides brasiliensis in tissue samples by a nested PCR assay

R Bialek et al. J Clin Microbiol. 2000 Aug.

Abstract

A nested PCR assay for the detection of Paracoccidioides brasiliensis DNA was evaluated, using a sequence of the immunogenic gp43 gene as a target. This gene encodes an outer membrane protein unique to this dimorphic fungus. DNA from six clinical isolates and the ATCC strain 60885 of P. brasiliensis, as well as DNA from closely related fungi, was examined to determine detection limits and cross-reactions. PCR was done on DNA extracts of lung homogenates from 23 experimentally P. brasiliensis-infected and two uninfected BALB/c mice and from 20 Histoplasma capsulatum-infected ICR mice. The results were compared to quantitative cultures. A detection limit of 0.5 fg of specific DNA was determined using cloned plasmid DNA. In all seven P. brasiliensis isolates, the expected 196-bp nested PCR product was found. Their sequences were 100% identical to the gp43 gene sequence in GenBank. DNA extracts of all other, related fungi were negative. The PCR assay was positive in 21 out of 23 culture-positive lung homogenates with concentrations of 1 x 10(3) to 1.3 x 10(7) CFU of P. brasiliensis per g of lung. Uninfected BALB/c mice and H. capsulatum-infected mice samples gave negative results. The high sensitivity and specificity of this new nested PCR assay for the detection of P. brasiliensis in tissue samples were demonstrated. The assay may be useful for diagnosis in human tissue samples.

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References

    1. Cisalpino P S, Puccia R, Yamauchi L M, Cano M I, da Silveira J F, Travassos L R. Cloning, characterisation, and epitope expression of the major diagnostic antigen of Paracoccidioides brasiliensis. J Biol Chem. 1996;271:4553–4560. - PubMed
    1. da Silva S P, Borges-Walmsley M I, Pereira I S, Soares C M D, Walmsley A R, Felipe M S S. Differential expression of an hsp70 gene during transition from the mycelial to the infective yeast form of the human pathogenic fungus Paracoccidioides brasiliensis. Mol Microbiol. 1999;31:1039–1050. - PubMed
    1. Franco L, Najvar L, Gómez B L, Restrepo S, Graybill J R, Restrepo A. Experimental pulmonary fibrosis induced by Paracoccidioides brasiliensis conidia: measurement of local host responses. Am J Trop Med Hyg. 1998;58:424–430. - PubMed
    1. Goldani L Z, Maia A L, Sugar A M. Cloning and nucleotide sequence of a specific DNA fragment from Paracoccidioides brasiliensis. J Clin Microbiol. 1995;33:1652–1654. - PMC - PubMed
    1. Goldani L Z, Sugar A M. Short report: use of the polymerase chain reaction to detect Paracoccidioides brasiliensis in murine paracoccidioidomycosis. Am J Trop Med Hyg. 1998;58:152–153. - PubMed

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